KR101499084B1 - Metal oxide scaffolds - Google Patents

Abstract

The present invention relates to metal oxide scaffolds comprising titanium oxide. The scaffolds of the present invention are useful for implantation into a subject to provide a framework for cell growth and stabilization for tissue regeneration and tissue regeneration. The present invention also relates to a method for producing such a metal oxide scaffold and its use.

Titanium, metal, oxide, scaffold, tissue regeneration, cell growth, osseointegration, implant.

Description

METAL OXIDE SCAFFOLDS < RTI ID = 0.0 >

The present invention relates to the field of implants comprising a material having improved properties in terms of mechanical stability and biocompatibility for implantation in a subject. Specifically, the present invention relates to an implant comprising a porous ceramic metallic material, and relates to a method for producing such a material for use as a bone conduction and osseointegration material.

Diseases such as trauma, tumors, cancer, periodontitis and osteoporosis can lead to bone loss, reduced bone growth and volume. For these and other reasons, it is very important to find ways to improve bone growth and restore anatomy. The scaffold may be used as a framework for cells participating in the bone regeneration process, and also as a framework for replacement of missing bone structures. What is also of interest is to provide a scaffold to be implanted in a patient having a surface structure that stimulates bone cells to grow to permit coating of the bone grafted structure after the healing process.

Orthopedic implants are used to preserve and restore function in the musculoskeletal system, particularly joints and bones, including relief of pain in these structures. A vascular stent is a tubular implant placed for insertion into a blood vessel to prevent or antagonize localized blood flow stenosis. In other words, they antagonize a significant reduction in blood vessel diameter.

Orthopedic implants are typically composed of materials that are stable in biological environments and that have minimal strain and endure physical stress. These materials must have strength, corrosion resistance, good biocompatibility, and good abrasion properties. Materials meeting these requirements include biocompatible materials such as titanium and cobalt-chromium alloys.

For tissue engineering purposes, it has been previously known to use a scaffold to support cell growth. Scaffold pore size, porosity and interconnectivity are thought to be important factors affecting cell function and regenerated cell quality. Prior art scaffolds are typically made of calcium phosphate, hydroxyapatite, and other types of polymers.

One principle of tissue engineering is to collect cells and, if necessary, expand the cell populations in vitro and seed them into a three-dimensional scaffold that supports the cells to grow into complete tissue or organs [1-5]. For most clinical applications, selection of scaffold materials and structures is important [6-8]. To achieve high cell density in the scaffold, the material needs to have a high surface area to volume ratio. The pores should be open and large enough to allow the cells to move to the scaffold. There must be sufficient space and passage to permit nutrient delivery, waste removal, material or cell depletion, and protein transport when cells are attached to the surface of the material, which can only be achieved with interconnected networks of pores [9, 10]. Biological responses to grafted scaffolds are also influenced by scaffold design factors such as three-dimensional microarchitecture [11]. In addition to the structural properties of the material, the physical properties of the material surface for cell attachment are essential.

Titanium and titanium alloys are frequently used as implant materials in dental and orthopedic surgery due to their biocompatibility with their bony tissue and their tendency to form a strong bond directly with bone tissue. This interaction between the bone tissue and the metal leading to this rigid attachment is called "osseointegration. &Quot;

Some metals or alloys, such as titanium, zirconium, hafnium, tantalum, niobium, or alloys thereof, that are useful for use in bone implants may be as strong as the bone tissue itself, or may have a relatively strong bond with bone tissue, Can be formed.

It is often desirable to improve this bond, although the bond between the metal, for example, titanium and the bone tissue, is relatively strong.

To date there are several ways to treat metal implants to obtain better adhesion of implants and thus improved bone adhesion. Some of these involve, for example, altering the shape of the implant by promoting a relatively large irregularity on the implant surface to increase the surface roughness compared to the untreated surface. Increased surface roughness provides greater contact and attachment area between the implant and bone tissue, thereby achieving good mechanical retention and strength. The surface roughness can be provided by, for example, plasma spraying, blasting or etching. The rough etching of the implant surface may be performed with a reducing acid such as hydrofluoric acid (HF) or a mixture of hydrochloric acid (HCl) and sulfuric acid (H ₂ SO ₄ ). The purpose of such a coarse etching process is to obtain an implant surface with rather large irregularities such as pore diameters in the range of 2-10 mu m and pore depths in the range of 1-5 mu m.

Other methods involve the modification of the chemical properties of the implant surface. This may also be done by modifying the surface nanostructure on the same basis with surface chemistry, for example by using a low concentration of fluoride solution, for example HF or NaF. For example, one such method involves, inter alia, the application of a layer of ceramic material, such as hydroxyapatite, to the implant surface to stimulate regeneration of the bone tissue. However, ceramic coatings are fragile and may peel or fall off the implant surface, which in turn may lead to ultimate failure of the implant.

It will also be noted that, in addition to the above-described methods of implant surface modification, titanium, zirconium, hafnium, tantalum, niobium, and alloys thereof are immediately covered with a thin oxide layer in contact with oxygen. The oxide layer of the titanium implant is mainly titanium dioxide (IV) (TiO ₂ ) and contains small amounts of Ti ₂ O ₃ and TiO ₂ . Titanium oxide generally has a thickness of about 4-8 nm.

WO 95/17217 and WO 94/13334 describe other methods of treating metal implants with an aqueous solution comprising fluoride. These two prior art applications describe metal implants with improved biocompatibility and methods of making such metal implants. Specifically, the rate of attachment of bone tissue is increased and a stronger bond between the implant and bone tissue is obtained. The improved biocompatibility of these implants is believed to be due to the retention of fluorine and / or fluoride on the implant surface.

Fluoride and / or fluoride can be prepared by the method of JE Ellingsen, "Treatment of titanium implants with fluoride improves retention of titanium implants in bone ", Journal of Material Science: Materials in Medicine, 6 (1995) 749-753), it is assumed that it reacts with the surface titanium oxide layer and replaces titanium-bonded oxygen to form a titanium fluoride compound. In vivo, the oxygen of the phosphate in the tissue fluid may replace the fluoride in the oxide layer and then be covalently bound to the titanium surface. This may lead to bone formation in which bone phosphates bind to titanium implants. Moreover, the released fluoride catalyzes this reaction and induces the formation of fluorinated hydroxyapatite and fluoroperatite in the surrounding bone.

WO 04/008983 and WO 04/008984 disclose other methods of improving the biocompatibility of implants. WO 04/008983 provides fluorine and / or fluoride on the implant surface and provides fine roughness to the surface with a mean square root roughness (Rq and / or Sq) of < 250 nm and / or a pore diameter < And pores having a pore depth of < = 500 nm. WO 04/008984 discloses a metal implant surface having a pore diameter of ≤ 1 μm and a pore depth of ≤ 500 nm and providing micro-roughness with pores having a peak width at half the pore depth of 15 to 150% The method comprising:

Bone ingrowth is preferential in a highly porous open cell structure, with cell size approximately equal to the cell size (0.25-0.5 mm) of the trabecular bone with struts with diameters of approximately 100 μm (0.1 mm) Is known to occur. Materials with high porosity with controlled microstructure are of interest to both orthopedic and dental implant manufacturers. For the orthopedic market, bone internal and external growth options currently include: (a) DePuy Inc. sintered metal beads on the implant surface to determine the appropriate pore size for intra-osseous growth, Resulting in a controlled microstructure with a lower porosity than the optimal porosity for the < RTI ID = 0.0 > (b) Zimmer Inc. uses fiber metal pads made by spreading loose fibers, the pads being either diffusion bonded to the implant or injection molded into the composite structure, which is also optimized for bone internal growth Has lower porosity than porosity; (c) Biomet Inc. uses a plasma sprayed surface, which results in a roughened surface that grows upwards and does not lead to intra-osseous growth; And (d) Implex Corporation uses a chemical vapor deposition process to produce tantalum-coated carbon microstructures, also called metal foams. Studies have suggested that this "trabecular metal" leads to high quality bone internal growth. The stranded metal has advantages of high porosity, open cell structure and cell size to aid bone internal growth. However, the support metal has a chemical composition and a coating thickness that are difficult to control. The matrix metal is very expensive, mainly due to the material and process costs associated with chemical vapor deposition (CVD) and long processing times. Moreover, CVD requires the use of highly toxic chemicals that are disgusting for biomedical uses.

The scaffolds available today are often degradable, which means that they are degraded after implantation in the subject. Although this is desirable in some cases, it may also be a disadvantage since it also leads to the loss of the stabilization function of the implant itself in other cases. In addition, prior art scaffolds often trigger inflammatory reactions and cause infections. For example, animal-derived bone implant scaffolds may cause allergic reactions when implanted in another animal. Metal implants with passive oxides have good biocompatibility, which means that the above disadvantages may be overcome by the use of implants made of such materials. However, it has not previously been possible to produce metal oxide scaffolds containing titanium oxide with sufficiently high mechanical stability to be practically useful.

One of the most promising biocompatible materials in this regard has been demonstrated in previous studies as bioactive ceramics, TiO ₂ [25-28]. This material exhibited specific biocompatibility, at which time scaffolds were transplanted in rats for 55 weeks without any inflammatory reaction or encapsulation [28]. Little research has been done in the production of three-dimensional open pores of titanium dioxide [29, 30]. The purpose of this study was to produce ceramic foams with defined macro-, micro-, and nanostructures and to illustrate possible uses of ceramic foams as scaffolds for cell cultures.

Summary of the Invention

It is therefore an object of the present invention to overcome the above-mentioned disadvantages, i. E. To have a good biocompatibility and without adverse effect when implanted in a subject, allowing cell growth in a three dimensional scaffold and still making it practically useful as a stabilizing structure To provide a metal oxide scaffold for use as a medical implant for implantation into a subject having mechanical stability. In addition, it is an object of the present invention to make the scaffold have an improved state for osteogenic cells to have surface properties that result in rapid bone growth at the scaffold surface and subsequently in the interconnected network of bone trabeculae will be.

The stated purpose is achieved by providing a metal oxide scaffold made of a metal oxide comprising titanium oxide, the scaffold having a compressive strength of about 0.1 to 150 MPa. More preferably, the metal oxide scaffold of the present invention has a compressive strength of 5-15 MPa.

In a second aspect, the present invention provides a medical implant comprising such a metal oxide scaffold.

In yet another aspect,

a) preparing a slurry of a metal oxide comprising titanium oxide, the slurry optionally comprising fluoride ions and / or fluorine,

b) providing a slurry of step a) to the porous polymer structure,

c) allowing the slurry of step b) to solidify,

d) removing the porous polymeric structure from the solidified metal oxide slurry.

Still yet another aspect, the present invention relates to the use of a medical implant as disclosed herein for regeneration, healing, replacement and / or repair of tissue such as bone, cartilage, cementum and dental tissue.

The present invention also relates to the use of a granulated metal oxide scaffold as a bone filling material.

The present invention further relates to a method of regenerating, healing, replacing and / or repairing a tissue comprising implanting a metal oxide scaffold or medical implant of a medical implant of the invention in a patient in need thereof.

Because the metal oxide scaffolds of the present invention are made of metal oxides containing titanium oxide with good biocompatibility, they can be implanted in a subject without causing adverse effects such as allergic reactions. The scaffold of the present invention also has a beneficial effect on the regeneration of the tissue due to the materials and surface structures thereof making them. The metal oxide scaffolds of the present invention are further particularly suitable for use in medical implants having sufficient stability to provide a stabilizing function while not yet solidified. This stability can be achieved by using titanium oxide without contamination of the secondary and / or tertiary phosphates.

1 is a hot plate forming (HPM). The depression in the plate prevents planarization of the viscous body during the sharpening process. The top is freely shaped by a combination of internal vapor pressure and viscosity.

Figure 2 is a flow chart of a polymer sponge method.

Figure 3 is a plot of the metal oxide scaffold. This landing did not collapse due to too short sintering time (A). This figure shows that the three-side strut collapses to remove the first internal void (B).

In Fig. 4, the percentage of bone inside voided cavities in which the scaffold rests describes the amount of newly formed bone. The reference standard is the apparent sham that the empty cavity left empty. Titanium oxide scaffolds, titanium oxide scaffolds with low fluoride, and titanium oxide scaffolds with intermediate fluoride content all had significantly (p < 0.001) increased bone quality inside the scaffold. The error bars represent the mean standard deviation and the height of the bars represents the average of the measurements (* p <0.001, n = 13).

In Figure 5, the post bone thickness box plot shows the amount of newly formed bone inside the scaffold. Titanium oxide scaffold with low fluoride and titanium oxide scaffold with intermediate fluoride content both had significantly (p < 0.001) increased bone formation inside the scaffold. The error bars represent the mean standard deviation and the height of the bars represents the average of the measurements (* p <0.001, ** p <0.05, n = 12).

6 shows the EDX mixture ratio of 80 wt% TiO ₂ and 20 wt% Al ₂ O ₃ .

7 is a Vickers (Vickers) gyeongdoyi ₂ -Al ₂ O ₃ in the composite having TiO other mixture.

Figure 8 is a Vickers hardness of a mixture of ZrO ₂ and TiO ₂ composite.

9 is a light microscope image of the sample _{60 wt% TiO 2/40 wt} % ZrO 2. The agglomerates are visible (white area) and the matrix is visible in the gray-black area.

Justice

Ceramic in this context means an object in which an inorganic powder material is treated with heat to form a solidified structure.

As used herein, the term "metal oxide" relates to oxides of metals such as oxides of Ti, Zr, Hf, V, Nb, Ta and / or Al. Examples of suitable metal oxides in the present invention include TiO ₂ , Ti ₃ O, Ti ₂ O, Ti ₃ O ₂ , TiO ₂ , Ti ₂ O ₃ , Ti ₃ O ₅ , ZrO ₂ , tantalum oxides (eg TaO ₂ ) And Al ₂ O _3, or combinations thereof.

As used herein, "scaffold" refers to an open porous structure. The scaffold structure herein typically has a pore size of about 10-3000 [mu] m, preferably 20-2000 [mu] m, more preferably 30-1500 [mu] m, and even more preferably 30-700 [ . Preferably, the pore size is above 40 [mu] m and has interconnection pores of at least 20 [mu] m.

The pore diameter thickness means the pure pore diameter, ie, the net diameter of the pore (not including the surrounding wall).

Fractal dimensionality is a statistical amount that gives an indication of how fully filled the fractals are, as they narrow down to a finer scale. There are many concrete definitions of the fractal dimension, and none of them should be treated as universal. The value of 1 belongs to a straight line. The larger the number, the more complex the surface structure.

The overall porosity is defined herein as all the compartments in the body, not the material, for example, a space not occupied by a substance. The overall porosity includes both closed and open porosity.

The internal strut volume refers to the volume of the inner lumen of the strut.

Contamination of the secondary and / or tertiary phosphate is defined to be present when the slurry as disclosed herein does not react to the ideal IEP (e.g., if the IEP of TiO ₂ is higher than 1.7, titanium oxide particles There is pollution).

"Sintering" means a method of making an object from a powder by heating the material (below the melting point) until the particles adhere to each other. Sintering has traditionally been used in the manufacture of ceramic bodies and in applications such as powder metallurgy.

A "medical implant" as used herein refers to a device intended to be implanted in the body of a mammal, such as a vertebrate such as a human being. Implants may be used herein to replace anatomical structures and / or to restore function of the body. Examples of implants include, but are not limited to, dental implants and orthopedic implants. The term "orthopedic implant" is used herein to refer to the body of a mammal such as a vertebrate animal, particularly a human, for the preservation and restoration of musculoskeletal, particularly joint and bony functions (including relief of pain of these structures) Any device intended to be implanted is within its scope. In the text, the term "dental implant" includes any device intended to be implanted in the oral cavity of a vertebrate animal, particularly a mammal such as a human, during a tooth restoration procedure. Dental implants may also be referred to as dental prosthetic devices. Generally, dental implants consist of one or several implant components. For example, a dental implant typically includes a dental fixture coupled to a secondary component, such as an abutment, and / or a dental restoration, such as a crown, bridge, or denture. However, any device, such as a dental fixture intended for implantation, may be referred to as an implant alone, even if other components are connected thereto.

In the text, the term "subject" refers to any vertebrate animal such as birds, reptiles, mammals, primates, and humans.

The mechanical strength of the scaffold was determined by a compression test for a load cell of 200 N (Zwicki, Zwick / Roell, Ulm, Germany). The scaffold was pre-loaded with 2 N force. The compression rate was set at 100 mm / min. The test data is testXpert 2.0. DIN EN ISO 3386, 1998.

The present invention relates to metal oxide scaffolds having improved biocompatibility and mechanical stability useful in medical implants. The invention also relates to a method of making such a scaffold and its use.

Scaffold implants are used to regenerate tissues after loss of tissue volume, as compared to solid implants (e.g., dental implants and hip prostheses) that replace tissues primarily to restore function. Since most prior art scaffolds are degradable, they are degraded to the patient after implantation. This means that they have the function of acting as a framework for the growth of the regenerated tissue. However, in some cases it is desirable to have a scaffold that also provides a stabilization function in addition to functioning as a framework for cell growth. However, as discussed above, previously available scaffolds have problems in terms of low biocompatibility, for example causing allergic reactions. It was also not possible to fabricate a metal oxide scaffold containing titanium oxide, which was previously stable enough to provide a stabilizing function while still solid when implanted in a subject.

The present inventors have surprisingly found that not only is it necessary to allow regeneration of the tissue but also to ensure that the scaffold itself is retained in the subject and has a biocompatibility that is also good practically made that allows the regenerated tissue to provide mechanical stability We have found a solution to the problem of providing scaffolds made of materials.

This object is achieved by providing a metal oxide scaffold comprising a metal oxide comprising titanium oxide. It is desirable that titanium oxide is capable of producing a metal oxide scaffold containing titanium oxide because it has the ability to engraft not only bone but also angiogenesis.

As described above, it has not been possible to produce a scaffold containing titanium oxide with stability that has previously been suitable for use as a medical implant. However, the metal oxide scaffold containing the titanium oxide of the present invention has a mechanical strength of about 0.1-150 MPa, which is a strength that is well suited for use as a medical implant. In a preferred embodiment, the mechanical strength is about 5-15 MPa. The compressive strength of the scaffold of the present invention is measured by a generally known method as described above.

The invention also relates to metal oxide scaffolds comprising titanium oxide as disclosed herein for regeneration, healing, replacement and / or repair of tissues, particularly bone tissue.

Most commercially available titanium oxide powders have surface contamination of secondary and / or tertiary phosphates. If these phosphate contaminations are used to produce a titanium oxide scaffold that interferes with proper sintering of the metal oxide scaffold during the manufacture of the metal oxide scaffold, the resulting scaffold will therefore not have satisfactory mechanical strength.

However, the present inventors have surprisingly found that by using titanium oxide without contamination of the secondary and / or tertiary phosphate on the surface of the titanium oxide particles (i.e., containing less than 10 ppm of such contaminants as described below) Metal oxide scaffolds can be prepared. Titanium oxides without secondary and / or tertiary phosphate on the surface of the titanium oxide particles may be obtained by using titanium oxide powders (such as titanium oxide powders from Sachtleben) that already have no such contaminants. Alternatively, the titanium oxide powder containing phosphate contaminants may be washed, such as by washing with NaOH (e.g., 1 M) to remove phosphate contaminants.

Titanium oxide scaffolds made of titanium oxide without phosphate contaminants have not been previously produced. As a result, titanium oxide scaffolds having previously practically useful mechanical strength could not be produced. In the present application, the titanium oxide comprises less than 10 ppm secondary and / or tertiary phosphate contaminants. These titanium oxides are believed to be free of secondary and / or tertiary phosphate contaminants herein.

The strength of the metal oxide scaffolds disclosed herein may vary by varying the porosity of the scaffold. Decreasing porosity will increase the strength of the scaffold

Kim et al. (24) disclosed a scaffold of ZrO ₂ with a hydroxyapatite (HA) layer. These scaffolds have a combination of mechanical strength due to ZrO ₂ and a biocompatible surface due to the HA layer. However, when loading using a scaffold, in these scaffolds, the HA layer may be peeled off or worn during implantation, or cracked, torn and loose from the surface of the implant due to differences in mechanical properties between ZrO ₂ and HA It may be. These HA fragments form sequestered bodies, which effectively initiate this body reaction and interfere with osseointegration of the scaffold. Moreover, another disadvantage of these ZrO ₂ scaffolds is that the ZrO ₂ surface itself has little or no inherent bone conduction effect when incorrectly exposed in the HA layer, and further contributes to these scaffolds being poorly adherent to the bone . By comparison, titanium oxide is more osmotic than ZrO ₂ and there is no risk of loss of this osmotic material when loading the scaffold since titanium oxide is an integral part of the scaffold of the present invention.

A further advantage of the inventive scaffold over the prior art is reduced manufacturing costs and easier manufacturing.

The metal oxide scaffold is intended for implantation into a subject, i. E., A medical implant for a subject such as a mammalian subject, e.g. a human patient. The metal oxide scaffold implants of the present invention include porous structures with improved surface properties that enhance biocompatibility and stimulate cell growth and attachment of implants. The porous structure permits internal growth of cells in the scaffold, thereby permitting tissue regeneration. The large surface area of the metal oxide scaffold also facilitates the growth of cells into the structure and thereby facilitates attachment of the scaffold and regeneration of the tissue. Since the scaffold is made of a material with good biocompatibility in itself, adverse effects on the scaffold when implanted in a patient are reduced.

The present invention provides a macroporous scaffold comprising macropores and interconnects. Macro voids of the metal oxide scaffold of the present invention range from about 10-3000 [mu] m, preferably about 20-2000 [mu] m, more preferably about 30-1500 [mu] m and even more preferably 30-700 [mu] m. More preferably, the macropore diameter is above about 100 [mu] m. Most preferably, the macropore diameter is about 30-700 [mu] m. For bones, pore sizes of 30-100 μm are optimal. However, it is also important that the scaffold also allows for internal growth of large structures, such as blood vessels and struts. That is, it is important to have pores of about 100 μm or more. It is important that at least some of the pores of the inventive scaffold are interconnected.

The pore size can be controlled by selection of the structure used to make the scaffold, e. G. The number of times the structure is immersed in a slurry containing the metal oxide (the process is described elsewhere herein) and the choice of sponge . By varying the pore size, it is possible to affect the rate and size of cell growth in the scaffold, thus affecting the resulting tissue composition. In another preferred embodiment, the metal oxide scaffolds comprise interconnected or partially interconnected pores. This means that the pores have at least two open ends, not " dead end "or closed pores, allowing passage of nutrients and waste products in more than one direction. This reduces the risk of formation of necrotic tissue. The macroporous system preferably occupies at least 50% volume of the scaffold. The volume of macro and micro pores in the scaffold may vary depending on the function of the scaffold. If the treatment objective is to replace a large amount of bone structure and the scaffold can be kept unloaded during the healing time, the scaffold may be made of a macroporous system that occupies up to 90% of the total scaffold volume.

Preferably the metal oxide scaffold according to the invention has a total porosity of about 40-99%, preferably 70-90%.

Preferably, the metal oxide scaffold according to the present invention has a fractal dimension support of about 2.0-3.0, preferably about 2.2-2.3. The strut thickness affects the strength of the scaffold, and the thicker the strut in the scaffold, the stronger the scaffold.

Preferably, the metal oxide scaffold according to the present invention has an internal pore volume of about 0.001-3.0 mu m &lt; ³ &gt;, preferably about 0.8-1.2 mu m &lt; ^{3 &} gt ;. Lower volume and higher fractal numbers provide a stronger scaffold.

Those skilled in the art will appreciate that the surface of the inventive scaffold also has micro- and nano-level structures. The micro- and nanostructures may be modified due to the manufacturing conditions. The pores produced by the manufacturing process are in the micro level ranging from 1 to 10 μm. The nano-level pores are less than 1 탆 in diameter.

The scaffold structure herein has a combined micro and macro pore size of about 10-3000 [mu] m, preferably 20-2000 [mu] m, more preferably 30-1500 [mu] m and even more preferably 30-700 [mu] m. Preferably, the pore size is above about 40 microns and has interconnect pores of at least 20 microns.

As will be apparent from the discussion below, due to the manner in which scaffolds are made, the scaffold of the present invention has a hollow tubular structure that creates bone gathers where bone grows and interconnects. Cells grow both inside and outside these tubules.

The size and shape of the metal oxide scaffold are determined according to the intended use. The size and shape of the resulting scaffold may be varied by varying the size and shape of the porous structure used in making the scaffold (see below). The scaffold can therefore be easily customized for specific applications to specific subjects.

The present invention relates to metal oxide scaffolds comprising titanium oxide, which further comprises fluoride and / or fluorine. Such a scaffold is such that the titanium fluoride formed is more chemically stable (i.e., less soluble from the surface results in the formation of a more stable scaffold) than the titanium oxide itself. In addition, fluorination of scaffolds has the additional advantage of providing improved biocompatibility. This may be due to the fact that the fluoride is believed to react with titanium oxide. The in vivo phosphate from the tissue will in turn replace the fluoride and the phosphate will bind to the titanium in the scaffold. This may lead to bone formation in which the phosphate in the bone binds to titanium implants. Moreover, the released fluoride catalyzes this reaction and leads to the formation of fluorophosphate and fluorophoreite in the surrounding bone. Thus, one embodiment of the invention is a metal oxide scaffold comprising at least one surface that is at least partially covered with fluoride and / or fluorine.

In one embodiment, the metal oxide scaffold is treated with an aqueous solution comprising fluoride ions and / or fluorine. For this purpose, an aqueous solution preferably containing HF, NaF and / or CaF ₂ is used. Alternatively, the fluoride / fluorine may be provided via the gas phase and / or as a vapor. The concentration of fluoride and / or fluorine in the aqueous solution is about 0.001-2.0 wt%, preferably 0.05-1.0 wt%. The pH of the aqueous solution comprising fluoride / fluorine is preferably about pH 0-7, preferably pH 2-6, more preferably pH 2-4.

In another embodiment, the fluoride ion and / or fluorine is provided in the slurry used to produce the metal oxide scaffold. As such, the fluoride / fluorine is provided as an integral part of the metal oxide scaffold.

By fluorination of the metal oxide scaffolds, their further increased stability is achieved. Thus, it is possible with the present invention to provide a scaffold with the advantages of high biocompatibility combined with improved mechanical stability and strength, so that the lost bone volume can be replaced by the scaffold and new bone can be regenerated And then a new bone structure can be created.

The oxides of titanium used in the metal oxide scaffold of the present invention include one or more titanium oxides selected from TiO ₂ , Ti ₃ O, Ti ₂ O, Ti ₃ O ₂ , TiO, Ti ₂ O ₃ , or Ti ₃ O ₅ do. In one preferred embodiment, the titanium oxide in the metal oxide scaffold comprises TiO ₂ . In one preferred embodiment, the metal oxide is titanium oxide comprising at least one titanium oxide selected from Ti ₃ O, Ti ₂ O, Ti ₃ O ₂ , TiO, Ti ₂ O ₃ , or Ti ₃ O _{5 in} combination with TiO ₂ . Preferably, the metal oxide used for the metal oxide scaffold of the present invention is TiO _2, Ti ₃ O, Ti ₂ O, Ti ₃ O _2, TiO, Ti ₂ O _3, or Ti ₃ O one or more of titanium selected from a _five Oxide. In yet another preferred embodiment, the titanium oxide in the metal oxide scaffold is comprised of TiO ₂ .

In addition to the titanium oxide, the metal oxide scaffold may further comprise one or more of oxides of Zr, Hf, V, Nb, Ta and / or Al as a mixture.

The metal oxide scaffold according to the present invention preferably has a titanium oxide content of about 40-100%, more preferably 60-90%, by weight in the metal oxide present in the scaffold.

In another embodiment, the metal oxide scaffold comprises ZrO ₂ . In this embodiment, ZrO ₂ is preferably at least about 90% of the metal oxide.

In another embodiment, the metal oxide scaffold comprises ZrO ₂ in combination with one or more of MgO, CaO, or Y ₂ O ₃ . This will improve the mechanical properties of the scaffold.

Another embodiment relates to a metal oxide scaffold in which the metal oxide comprises Al ₂ O ₃ . In this embodiment, Al ₂ O ₃ is at least about 90% of the metal oxide.

In yet another embodiment of the present invention, the metal oxide scaffold comprises a complex of Al ₂ O ₃ and TiO ₂ . Complex may also comprise TiO ₂ and ZrO _2. These scaffolds will have increased strength.

The metal oxide scaffold according to the present invention may also comprise particles of metal oxide (about 10 nm-100 탆 in size) which are mixed with the metal oxide slurry during the production of the metal oxide scaffold. Scaffolds prepared in this way containing particles of metal oxide dispersed in metal oxide have increased stability.

The metal oxide scaffolds of the present invention may also optionally include CaPO ₄ , Cl ^- , F ^- and / or carbonates. Such additives may also increase the biocompatibility of the scaffold and improve their osseointegration.

In yet another aspect, the present invention relates to a medical implant comprising a metal oxide scaffold. The medical implant according to the present invention may be the metal oxide scaffold itself in some embodiments of the present invention. Alternatively, the medical implant may comprise a metal oxide scaffold of the present invention in combination with an orthopedic, dental or any other fixture or other structure such as an implant. The invention also relates to medical implants comprising a metal oxide scaffold comprising titanium oxide for the regeneration, healing, replacement and / or repair of tissue, particularly bone tissue.

In another aspect, the invention relates to a slurry for preparing a metal oxide scaffold comprising titanium oxide. The slurry comprises an aqueous solution of a titanium oxide powder, wherein the titanium oxide powder comprises less than about 10 ppm of secondary and / or tertiary phosphate contaminants. The aqueous solution contains ordinary water or deionized water as a solvent. The ratio of titanium oxide to moisture content (weight ratio) is at least 2: 1. The slurry may also comprise other metal oxides as described herein. Optionally, the slurry may also contain fluoride ions or fluorides (e.g., at a concentration of 0.01 wt% HF). Alternatively, the slurry may also contain other components such as CaPO ₄ , Cl ^- , F ^- , carbonate, MgO, CaO, and / or Y ₂ O ₃ , which are interesting in the metal oxide scaffold. The slurry is preferably an aqueous slurry and withstands a temperature of at least 500 &lt; 0 &gt; C.

The titanium oxide powder used to make the slurry may be amorphous, anatase, brucite or rutile crystalline phase.

The particle size of the metal oxide in the slurry is advantageously as small as possible to ensure proper sintering (see below for details of the sintering process). Thus, metal oxide particles need to be reduced to smaller pieces and need to be evenly distributed prior to sintering. By using titanium oxide containing less than about 10 ppm of secondary and / or tertiary phosphate in the preparation of the slurry, the titanium oxide particles are sufficiently small to permit proper sintering without the addition of an organic anti-aggregation compound. This leads to an easier manufacturing process. Therefore, preferred embodiments of the present invention relate to slurries comprising no additives such as organic additives and / or titanium oxide to which no surfactant is added. However, if other metal oxides other than dithium oxide are present in the scaffold, a binder such as a polysaccharide binder (e.g. Product KB 1013, Zschimmer & Schwarz GmbH, Lahnstein, Germany) may be added to reduce the aggregation and stabilize the slurry. May be required to add an organic additive such as a &lt; RTI ID = 0.0 &gt;

Typical slurries used in the present invention include water, metal oxide particles and optionally organic parts such as binders and / or surfactants for reducing surface tension.

One preferred embodiment relates to metal oxide scaffolds comprising TiO ₂ with less than about 10 ppm secondary and / or tertiary phosphate. Another preferred embodiment relates to a metal oxide scaffold containing only TiO ₂ with less than about 10 ppm secondary and / or tertiary phosphate as metal oxide in the scaffold.

The slurry for preparing the metal oxide scaffold containing titanium oxide has a pH value of about 1.0 to 4.0, preferably about 1.5 to 2.0. It is desirable to reduce the size of the titanium oxide particles as close as possible to the pH value that provides the theoretical isoelectric point of the titanium oxide. The pH value for TiO ₂ is 1.7. The average particle size of the titanium oxide particles is preferably 10 占 퐉 or less, more preferably 1.4 占 퐉 or less. Preferably monodisperse.

In preparing the metal oxide scaffold of the present invention, the starting slurry comprising the metal oxide is aqueous and it is important to withstand the combustion at temperatures above 500 ° C.

The method for producing a metal oxide scaffold of the present invention comprises

a) preparing a slurry of a metal oxide comprising titanium oxide, the slurry optionally comprising fluoride ions and / or fluorine,

b) providing a slurry of step a) to the porous polymer structure,

c) allowing the slurry of step b) to solidify,

d) removing the porous polymeric structure from the solidified metal oxide slurry.

Details of the composition of the slurry used in this process are described elsewhere herein.

The slurry is prepared by dispersing the metal oxide and other optional components in the solvent used. Preferably, the metal oxide is gradually added to the solvent while stirring and reconditioning the pH value, for example with 1 M HCl, to maintain the desired pH value. The slurry is then further dispersed, for example, into a rotating nanodisperse (dispermat) having metal blades, preferably titanium blades. Preferably this is carried out for at least 4 hours at a rate of at least 4000 rpm. More preferably, this step is carried out at 5000 rpm for at least 5 hours. The pH of the slurry is regularly adjusted to the selected pH value.

The slurry is then provided to the porous polymeric structure. The porous polymer structure may be, for example, a sponge structure such as a synthetic sponge. The material is preferably made of an organic material to facilitate the removal of the porous polymer structure from the scaffold by combustion. Therefore, the porous polymer structure is an organic sponge structure, preferably an organic porous polymer sponge, preferably a polyethylene, silicone, cellulose or polyvinyl chloride sponge. One example of a preferred porous polymeric structure is a 45 or 60 ppi Bulbren polyurethane foam (Bulbren S, Eurofoam GmbH, Wiesbaden, Germany). The porous polymeric structure is preferably washed with water to remove residues and / or contaminants prior to providing the slurry thereto. The slurry may be provided to the porous polymeric structure by dipping the porous polymeric structure into a slurry. The excess slurry after dipping can be removed by squeezing and / or centrifuging the porous polymer structure immersed in the slurry. The porous structure dipped in the slurry is then dried, for example, for at least 5 hours and more preferably for about 24 hours to allow the slurry to solidify in the porous polymer structure. By varying the number of times to perform step b) -d), the size of the macropores of the scaffold can be varied.

The size and shape of the metal oxide scaffolds produced can be adjusted by adjusting the size and shape of the porous polymer structure used. Thereby making it possible to produce customized scaffolds for the specific intended implantation site of a particular patient.

The next step in the preparation of the metal oxide scaffold is to remove the porous polymeric structure therefrom from the solidified slurry to obtain a scaffold structure.

In a preferred method of the invention, step d) is carried out by removing the porous polymeric structure from the solidified metal oxide by heating.

Preferably, the porous polymer structure is a combustible structure. Thus, step d) in the above method may be carried out, for example, by burning the porous polymer structure from the solidified metal oxide slurry. The temperature and time required to perform this process depends on the material from which the porous polymer structure is made, as will be readily appreciated by those skilled in the art. Importantly, the acceptable temperature and time must be selected for a rather complete removal of the porous polymer structure. One skilled in the art will know how to select the time and temperature required for a particular porous polymer structure and scaffold to achieve this.

In a preferred method step d)

i) slowly sintering the porous polymeric structure with the solidified metal oxide slurry to about 500 DEG C and holding the temperature for at least 30 minutes

ii) rapidly sintering at about 3 K / min from about 1500 [deg.] C to about 1750 [deg.] C, holding the temperature for at least 10 minutes

iii) rapid cooling to room temperature at least 3 K / min.

The biological response to the metal oxide scaffold can be varied in different ways. One example is to vary the crystalline phase of the coated layer of titanium oxide. The crystalline phase can be changed by terminating the sintering of the coated layer at different temperatures. Sintering above 960 ± 20 ° C provides a rutile phase while sintering below 915 ± 20 ° C provides an anatase phase.

The surface of the resulting scaffold may be modified, for example, by providing a fluoride ion and / or fluoride thereon. The fluoride and / or fluorine may be provided in an aqueous solution comprising HF, NaF and / or CaF ₂ , wherein the fluoride and / or fluorine in the solution comprises about 0.001-2.0 wt%, preferably 0.05-1 wt% And the fluoride and / or fluorine is provided to the surface of the scaffold by immersing the scaffold in the solution. Ideally, this is not done in excess of 120 seconds with a solution having 0.1 wt% fluoride and / or fluorine. Alternatively, the fluoride / fluorine may be provided via the gas phase and / or as a vapor.

In addition, biomolecules may be provided on the surface of the scaffold. If biomolecules are provided to the metal oxide scaffold, they may be provided after step d) of the method. The presence of biomolecules may also increase the biocompatibility of the scaffold and the rate of cell growth and attachment. Biomolecules are used herein to refer to natural biomolecules (i.e., naturally occurring molecules derived from natural sources), synthetic biomolecules (i.e., naturally occurring biomolecules prepared by synthesis, and forms of non-naturally occurring molecules or molecules produced by synthesis ) Or recombinant biomolecules (prepared through the use of recombinant techniques). Examples of interesting biomolecules include but are not limited to bioadhesives, cell attachment factors, biopolymers, blood proteins, enzymes, extracellular matrix proteins and biomolecules, growth factors and hormones, nucleic acids (DNA and RNA), receptors, synthetic biomolecules, But are not limited to, biomolecules disclosed in US2006 / 0155384 such as biologically active ion marker biomolecules including proteins and peptides such as drugs, statins, and proteins or peptides that stimulate bio-mineralization and bone formation. The biomolecule can be attached to the surface of the scaffold, for example, by immersion in a solution containing biomolecules or by an electrochemical process. Such processes are well known to those skilled in the art.

In another aspect, the present invention relates to a metal oxide scaffold as disclosed herein for use as a medical implant. One embodiment relates to a medical implant for use for tissue regeneration. Another embodiment relates to a medical implant for use for bone regeneration.

In another aspect, the invention also relates to the use of a metal oxide scaffold as disclosed herein for the manufacture of a medical implant for regeneration, healing, replacement and / or repair of tissue such as bone tissue.

The metal oxide scaffold of the present invention may also be granulated or used as a bone filling material. The advantage of using a granulated scaffold as a bone filling material is that it may be used to fill a bone void (boneless pocket, where the scaffold can be granulated to fill all the empty volume). Therefore, in yet another aspect, the present invention relates to a granulated bone filling material comprising a metal oxide scaffold according to the present invention. For this purpose, the ground metal oxide scaffold particles may be classified by particle size. In order to fill the bone cavity, it is optimal that the particle size is in the range 0.05-5 mm (mean diameter), preferably 0.1-2 mm, and most preferably 0.2-1 mm. A mixture of particles of the same size or of different sizes may be used.

In yet another aspect, the invention relates to a metal scaffold as disclosed herein granulated for use as a bone filler material. In still yet another embodiment, the present invention relates to the use of a metal scaffold as disclosed herein, granulated in the preparation of a bone filler material.

Still yet another aspect, the present invention also relates to a method for regeneration, healing, substitution and / or repair of tissue comprising implanting a metal oxide scaffold or a medical implant into a subject in need thereof.

Tissue engineering involves the development of new generations of biomaterials capable of specific interactions with biological tissues to produce functional tissue substitutes. The underlying idea is that cells can be isolated from patients, expanded in cell cultures and seeded into scaffolds made from specific biomaterials to form a scaffold / biological complex called the "TE structure ". The structure may then be grafted to the same patient and function as an alternative tissue. Some of these systems are useful for replacing organ tissues where there is limited availability of donor or, in some cases (eg, young patients), where inappropriate natural substitutes are available. The scaffold itself may act as a delivery vehicle for growth factors, genes and biologically active portions from the drug. This innovative approach to surgery has a wide range of applications that benefit both patient well-being and advances in healthcare systems.

The scaffold of the present invention can be implanted into cells and cells will grow in the scaffold construct. It is also possible to seed and grow the cells in the implant prior to implantation. The novel interconnected macroporous structures of the metal oxide scaffolds of the present invention are particularly well suited to tissue engineering and are eminently suitable for bone tissue engineering and are an interesting alternative to currently available bone healing therapies. In this regard, bone marrow-derived cell seeding of the metal oxide scaffold is carried out using conventional methods well known to those skilled in the art (Maniatopoulos et al, in Cell Tissue Res 254, 317-330, 1988). The cells are seeded in a metal oxide scaffold and cultured under suitable growth conditions. The cultures are grown in media suitable for promoting growth.

As indicated above, various types of cells can be grown through the metal oxide scaffold of the present invention. More precisely, cell types include hematopoietic or mesenchymal stem cells, and also include cells that produce cardiovascular, muscular, or any connective tissue. The cells may be derived from humans or other animals. However, the metal oxide scaffolds of the present invention are particularly suitable for the growth of osteogenic cells, particularly cells producing bone matrix. For tissue engineering, a cell can be of any origin. Cells are of human origin. The method of the invention for growing cells in a three-dimensional metal oxide scaffold according to the present invention is characterized in that seeded osteogenic cells penetrate the metal oxide scaffold, for example, and have an invasive distribution in the structure of the metal oxide scaffold, Allows the creation of bone matrix during the in vitro stage. Penetration of osteogenic cells and, as a result, bone matrix formation can be enhanced by mechanical, ultrasonic, electric or electronic means.

The scaffold of the present invention is useful whenever there is a need for a structure that acts as a framework for cell growth, such as for tissue regeneration. The scaffold of the present invention is particularly useful for the regeneration of bone and cartilage structures. Examples of situations in which regeneration of such structures may be required include surgical removal of traumas, bones or teeth, or those associated with cancer therapy. Therefore, from a different viewpoint, the present invention also relates to the use of medical implants as disclosed herein for regeneration of tissues such as bone, cartilage, cementum, and tooth tissue, and the use of such tissues As well as to a method for reproducing the same.

Examples of structures in a patient that may be replaced in whole or in part include: head-face bone including cheekbones, bone of inner ear (especially hammer bone, crus bone and ankle), jaw and lower teeth tooth, wall and bottom of orbit, (Including but not limited to) the walls and floor of the oysters, defects in the skull and skull, fossa acetabuli, such as in the case of hip arthroplasty, humerus, radial, ulnar, femur, But are not limited to, multiple fractures of the bones, vertebral bones, bone of the hands and feet, bones of the fingers and toes, filling of the extraction shaft (from tooth extraction), healing of periodontal defects and healing of implants.

In addition, the scaffolds of the present invention are useful for filling all types of bone defects resulting from tumors, infections, trauma, surgery, congenital malformations, genetic diseases, metabolic diseases (e.g., osteoporosis and diabetes).

Example 1: Preparation of TIO ₂ slurry

The slurry used consisted of an electrostatically stabilized TiO ₂ - suspension with no further additives. The components of the slurry are deionized water, TiO ₂ - powder (batch 1170117, Sachtleben Hombitan Anatase FF-Pharma, Duisburg, Germany) and 1 mol / l HCl (Merck Titrisol, Oslo, Norway). The suspension was set to pH 2.2 by HCI. To achieve optimum dispersion, a suspension was prepared in a stirrer mill with a ceramic double grinding disk. A 600 g Zirconox CE milling bead (Jyoti GmBH, Drebber, Germany) (having a diameter of 0.4-0.7 mm) was used as the grinding body. The preparation of the slurry was carried out in the following manner:

1. Place 118.8 g of deionized water with 1.2 g of 1 mol / l HCl in a water-cooled container with milling beads.

2. The addition of the total amount of 210 g TiO ₂ - powder occurs gradually. Gradually added to the liquid TiO ₂ -Pulver a stirring speed of about 1000 RPM. The viscosity of the suspension gradually rises. This action is due to the fact that the protons from the liquid reservoir attach themselves to the OH groups of TiO _{2 in} the powder. The pH value in the suspension therefore shifts to pH 4.2. To lower the viscosity again, titrate 1 M HF in the suspension with a pipette. Usually 1 ml HF is sufficient. The viscosity is thus adjusted with additional HCl in the slurry.

3. The HCl titration continues until the total amount of 210 g TiO ₂ is completely dispersed. About this amount, about 8 ml of 1 M HCl are usually needed. Thus, the solids content in the suspension is about 30 vol. %.

4. After this dispersion, the agglomerates and agglomerates are destroyed by milling at a high speed of 4000 RPM. Optimum results are obtained after approximately 5-6 hours.

5. The viscosity of the suspension can be strongly influenced by the purpose of use by varying the solids content or pH value. Higher viscosity is intended for the manufacture of thicker scaffolds, while thinner and finer structures require lower viscosity.

6. Small bubbles may interfere with the production of ceramics and thus must be defeated. This can be done, for example, by rotary evaporation or by an ultrasonic bath. In addition, the slurry can be filtered with a mesh of pore sizes of 100 [mu] m and 50 [mu] m. This process also reduces heterogeneity.

7. There are several ways to make a scaffold after obtaining a suitable ceramic slurry. The following two sections describe two methods, the hot plate forming reported by Eckardt et al [12] and the polymer sponge method described by Haugen et al [13].

Example 2: TiO ₂ slurry preparation with fluoride doping

The slurry used consisted of an electrostatically stabilized TiO ₂ - suspension with no further additives. The components of the slurry are deionized water, TiO ₂ - powder (batch 1170117, Sachtleben Hombitan Anatase FF-Pharma, Duisburg, Germany) and 1 mol / l HCl (Merck Titrisol, Oslo, Norway). The suspension was set to pH 2.2 by HCI. To achieve optimum dispersion, a suspension was prepared in a stirrer mill with a ceramic double grinding disk. A 600 g Zirconox CE milling bead (Jyoti GmBH, Drebber, Germany) (having a diameter of 0.4-0.7 mm) was used as the grinding body. The preparation of the slurry was carried out in the following manner:

1. Place 118.8 g of deionized water with 1.2 g of 1 mol / l HCl in a water-cooled container with milling beads.

2. The addition of the total amount of 210 g TiO ₂ - powder occurs gradually. The TiO ₂ powder is added stepwise to the liquid at a stirring rate of about 1000 RPM. The viscosity of the suspension gradually rises. This action is due to the fact that the protons from the liquid reservoir attach themselves to the OH groups of TiO _{2 in} the powder. The pH value in the suspension therefore shifts to pH 4.2. To lower the viscosity again, titrate 1 M HF in the suspension with a pipette. Usually 1 ml HF is sufficient. The viscosity is thus adjusted with additional HCl in the slurry.

3. Adding HF acid to the slurry also has the effect of doping TiO ₂ into the titanium-fluoride containing compound.

4. The HF titration continues until the total amount of 210 g TiO ₂ is completely dispersed. About this amount, about 8 ml of 1 M HF is usually needed. Thus, the solids content in the suspension is about 30 vol. %.

5. The amount of doping is controlled by the amount of HF added to the slurry and can be replaced with 1 M HCl to lower the fluoride concentration.

6. After this dispersion, the agglomerates and agglomerates are destroyed by milling at a high speed of 4000 RPM. Optimum results are obtained after approximately 5-6 hours.

7. The viscosity of the suspension can be strongly influenced by the purpose of use by varying the solids content or pH value. Higher viscosity is intended for the manufacture of thicker scaffolds, while thinner, finer structures require lower viscosity.

8. Small bubbles may interfere with ceramic manufacture and thus must be defeated. This can be done, for example, by rotary evaporation or by an ultrasonic bath. In addition, the slurry can be filtered with a mesh of pore sizes of 100 [mu] m and 50 [mu] m. This process also reduces heterogeneity.

Example 3: Scaffold Fabrication - Hot Plate Forming

Hot plate forming is a shaping method for shaping suspension droplets containing a ceramic raw material into an empty spherical geometry using the effect of water vapor (Fig. 1). A fluoride doped TiO ₂ slurry was prepared (see Example 2) and mixed with 6.0 g of a polyethylene-copolymer powder (Terpolymer 3580, Plastlabor, Switzerland) and 5.0 g of phenolic resin powder (FP 226, Bakelite, Germany). The brass plate with spherical recesses was heated to 320 ° C. Slurry drops were then provided to each well using a 2 ml syringe. Heat transfer from the plate to the droplets resulted in the emission of water vapor, which then formed bubbles within the droplets. The agglomeration of these water droplets created a large cavity in the center of the droplet and prevented the structure from collapsing with progressive drying. The continuously rising water vapor pressure eventually resulted in the rupture of the topmost part of the droplet still. Surface tension acts to round the rim of this opening. Heat transfer from the hot plate also melted and pyrolyzed the polymer powder in the formed body, thus stabilizing the vacant sphere shape and separating it from the hot plate. The polymers were removed by heating to &lt; RTI ID = 0.0 &gt; 800 C &lt; / RTI &gt; The pre-fired samples were sintered in an electric furnace at 1620 ° C for 15 minutes and brought into a porous ceramic [12].

Example 4: Preparation of Scaffold-Polymer Sponge Method

Open-pore ceramics are produced by the replication of polymer porous structures. The patent for this technique, called the "cloning" or "polymer-sponge" method, was first filed in 1963 by Schwartzwalder and Somers [14]. This is the standard method for making alumina, zirconium, silicon carbide and other ceramic foams [15-19]. Foams are prepared by coating the polyurethane foam with the prepared fluoride-doped TiO ₂ slurry (see Example 2). Polymers already having the desired macrostructure are simply provided as sacrificial scaffolds for ceramic coatings. The slurry infiltrates the structure and adheres to the surface of the polymer. Excess slurry is squeezed to leave a ceramic coating on the foam strut. After drying, the polymer is slowly burned to minimize damage to the porous coating. Once the polymer is removed, the ceramic is sintered to the desired density. The process duplicates the macroscopic structure of the polymer and produces a rather unique microstructure within the strut. A flow diagram of the process was provided (Figure 2).

Example 5: Scaffold production - Polymer sponge method with coating

Open-pore ceramics are produced by the replication of polymer porous structures. The patent for this technique, called the "cloning" or "polymer-sponge" method, was first filed in 1963 by Schwartzwalder and Somers [14]. This is the standard method for making alumina, zirconium, silicon carbide and other ceramic foams [15-19]. Foams are prepared by coating a polyurethane foam with the prepared TiO ₂ slurry (see Example 1). Polymers already having the desired macrostructure are simply provided as sacrificial scaffolds for ceramic coatings. The slurry infiltrates the structure and adheres to the surface of the polymer. Excess slurry is squeezed to leave a ceramic coating on the foam strut. After drying, the polymer is slowly burned to minimize damage to the porous coating. Once the polymer is removed, the ceramic is sintered to the desired density. The foam is now immersed in the fluoride-doped TiO ₂ slurry described in Example 2, followed by drying and sintering. The immersion process is continued until a uniform coating of fluoride-doped TiO ₂ covers the TiO ₂ scaffold.

Example 6: Slurry preparation

The slurry used consisted of an electrostatically stabilized TiO ₂ - suspension and no further organic additives. The components of the slurry are sterile water, TiO ₂ - powder (Pharma FP Hobitam, Sachtleben GmbH, Duisburg, Germany) and 1 mol / l HCl (Merck, Oslo, Norway). The suspension was set to pH 1.7 by HCI. To achieve the optimum particle size, the ceramic slurry was dispersed in a high speed dissolution apparatus (Dispermat Ca-40, VMA-Getzmann GmbH, Reichshof, Germany) with a custom titanium rotary blade. The preparation of the slurry was carried out in the following manner:

1. 72 grams of TiO ₂ powder was gradually added to 29.7 ml of deionized water at pH 1.7 in a water-cooled vessel under stirring at a low rotational speed of 1000 rpm.

2. The TiO ₂ powder was gradually added while maintaining the temperature at 20 ° C to 25 ° C. When the rotational speed was increased to 4000 rpm, the temperature was decreased to 10 ° C-18 ° C.

3. When the slurry was homogeneous, the rotational speed was increased to 5000 rpm for 5 hours.

4. The pH was adjusted every 30 min and readjusted to pH 1.7 with 1 mol / L HCl.

Example 7: Scaffold Fabrication - Polymer Sponge, Second Embodiment

Open-pore ceramics are produced by the replication of polymer porous structures. This method is called the "schwartzwald method" or the polymer sponge method [14]. A fully homogeneous polyester-based polyurethane foam (Bulbren S, Eurofoam GmbH, Wiesbaden, Germany) with 60 ppi was used in this study. The foams were fed into a large 8 mm thick plate and punched out with a metal stamp to cut into a cylinder of 12 mm diameter. The tablet was then washed for 2 minutes in 1 l deionized H ₂ O and 10 ml Deconex (Burer Chemie AG, Zuchwill, Switzerland) and then in ethanol (Absolute, Arcus, Oslo, Norway). The tablet was then dried at room temperature for 24 hours and stored in PE-bag. The slurry was prepared as described in Example 6. The polymer foam was then dipped into the ceramic slurry. These foams were then centrifuged at 1500 rpm for 2 minutes at 18 DEG C (Biofuge 22R Heraeus Sepatech, Osterode, Germany). The next sample was then placed on a porous ceramic plate prior to sintering and allowed to dry at room temperature for at least 24 hours. The heating schedule for the combustion of the polymer and the sintering of the ceramic parts was selected as follows: slowly heated to 450 DEG C at 0.5 K / min, held for 1 hour at 450 DEG C, heated to 1500 DEG C at 3 K / min, Holding time, cooling to room temperature with 6 K / min (HTC-08/16, Nabertherm GmbH, Bremen, Germany).

order Temperature
Range (℃) heating
speed
(K / min) duration
(min) One 25-450 0.5 850 2 450 0 60 3 450-1500 3 350 4 1500 0 3000 5 1500-50 6 242

Sintering process

When the maximum temperature of 1500 ° C was achieved and when this temperature was maintained for more than 20 hours, the collapse of the ceramic strut was reached. As the energy was transferred to the scaffold, the void space created by burning the polymer foam collapsed and a more dense and stronger structure was created (FIG. 3).

The sintering process produced a nanostructured surface on the fused TiO ₂ particles. You can see the grain boundaries where fusion occurs. At a high magnification, a wave-like nanostructured surface can be seen (not shown).

The structures on sintered titanium oxide particles had the following roughness parameters provided in Table 2:

Surface states and morphology of titanium oxide scaffolds (n = 5). Roughness parameters of titanium oxide scaffold surface applied to single particles Sa Sq Ssk Sku Sfd (1) Sci ㎛ ㎛ <No unit> <No unit> <No unit> <No unit> Average 0.89 1.12 -0.87 3.28 2.35 0.53 Standard Deviation 0.42 0.56 0.31 0.79 0.06 0.26

In the titanium oxide scaffold, the surface roughness, Sa, was 890 nm, and the mean square root, Sq, was 1.12 탆. Since the asymmetry, Ssk, is slightly negative, there are more surfaces with a valley than a peak. The sharpness, Sku, has been shown to have a steep peak. The number of fractures, Sfd, indicates the complexity of the surface, the fluid retention number, and Sci has a range of correlated values of bone attachment and volume.

Typically, Sa is 0.3-1.1 占 퐉, Sq is 0.4-1.4 占 퐉, Ssk is -1.2 to 1.2, Sku is 1-4 and Sci is 0.2-1.5 占 퐉. Sa is preferably about 0.9. Preferably, Sq is about 1.2. Preferably, Ssk is -0.9. Preferably Sku is 3. Preferably, Sci is 0.63.

Characterization of Titanium Oxide Scaffolds Doped with Fluoride Pore Structures

This structure is similar to the TiO ₂ scaffold described.

Example 8: Preparation of scaffolds - Polymer sponge, double coating

The scaffold was made and sintered as described in Example 7. [ These scaffolds were then immersed in the slurry as described in Example 6 and then centrifuged at 1500 rpm for 2 minutes at 18 DEG C (Biofuge 22R Heraeus Sepatech, Osterode, Germany). The samples were then placed in a porous ceramic plate And dried at room temperature for at least 24 hours prior to sintering The second sintering step was carried out at a rate of 3 K / min at 1500 ° C and at 1500 ° C for 30 hours.

Example 9 Scaffold Fabrication - Polymer Sponge, Doubled Coating Doped with Fluoride

A scaffold was prepared as described in Example 8. The scaffold was then immersed in 0.2 wt% hydrochloric acid. 20 scaffolds were immersed for 90 seconds and 20 scaffolds were immersed for 120 seconds. The scaffold was then rinsed in sterile water for 1 minute and air-dried in a layer flow and packed in sterile packaging.

Example 10 Characterization of Titanium Oxide Scaffolds Prepared According to Examples 8 and 9

(Skyscan, Aartselaar, Belgium) with a 0.7 m voxel resolution (50x magnification), X-ray tube current of 173 및 and a voltage of 60 kV, with a 0.5 mm aluminum filter, scaffold (n = 49) - line CT scanner. The specimen was mounted vertically on a plastic support and rotated 180 DEG about the long axis (z-axis) of the sample. The four absorbed images were recorded every 0.300 ° rotation. These projection random images were used for standard cone-beam reconstruction to generate a series of 1024 8-bit axial slices, each with 1024 x 1024 pixels, with Z-dimensional spacing such as the inter-slice pixel spacing. The resulting 3-D data set was therefore isotropic with a voxel spacing of 7 μm across the entire 1024 ³ spatial range. 3-D reconstructions of internal pore shapes were performed using these axial bitmap images and analyzed by CTan and CTvol (Skyscan, Aartselaar, Belgium). At this time, the gray scale threshold value was set to 55 to 230. Additional noise was removed by the "despeckling" function. Thus, all white objects smaller than 50 voxels were removed prior to further analysis. All the images were 3D analyzed and then "shrink-wrap", which allowed to measure the volume of the empty stanchions. The density of the scaffold column was measured using the same software, and the calibration was taken at 1.25 and 1.75 g / cm3. From the gray scale to the density, a Hounsfield unit calibration was performed. The relationship was linear.

The BET surface area was found to be 5.0664 m 2 / g when analyzed with liquid nitrogen (TriStar 3000, Micromeretics, Monchengladbach, Germany).

In vivo experiment

Three types of scaffolds were tested as described in Examples 8 and 9. A dual coated titanium oxide scaffold is referred to herein as TiO ₂ . A dual coated titanium oxide scaffold immersed in 0.2 wt.% HF for 90 seconds is referred to herein as TOS. A double coated titanium oxide scaffold dipped in 0.2 wt.% HF for 120 seconds is referred to herein as THF.

Animal and Surgery courses

Eighteen (18) New Zealand white rabbits aged 8 months and 3.0-3.5 kg were used in the study (ESF Produkter Estuna AB, Norrtalje, Sweden). The animals were placed on us during the trial period. The room temperature was adjusted to 19 ± 1 ℃ and the humidity was 55 ± 10%.

The experiment was approved and registered with the Norwegian Animal Research Authority (NARA). The procedure was therefore carried out in accordance with the Animal Welfare Act of December 20th 1974, No 73, Chapter VI, Sections 20-22 and Regulation on Animal Experimentation of January 15th 1996. Ten minutes before the rabbits were taken from us, they were injected with 0.05-0.1 ml / kg of subcutaneous fluianzone / fentanyl (Hypnorm®, Janssen, Belgium) and 2 mg / kg bw intravenously Midazolam (Dormicum®, Roche, Switzerland). When there were any signs of waking, diluted Hypnorm® was slowly injected intravenously until a suitable effect was achieved. Lidocain / adrenaline (Xylocain / Adrenalin®, AstraTech AB, Molndal, Sweden) 1.8-ml s.p. Before surgery, the surgical site was removed and cleaned with a soft soap. The animals were placed back on the table, covered with a sterile cloth, and the surgical site was disinfected with Chlorhexidingluconat 5 mg / ml (Klorhexidin, Galderma Nordic AB, Sweden).

An incision was made to penetrate all the soft tissues in the anterior tibia of the tibia. The periosteum was lifted and maintained by a self-retaining retractor. The drill guide was used to create four guide holes with a twist drill (Medicon® CMS, Germany) to ensure standardized correct positioning. A platform for titanium implants was made using a custom stainless steel bur (6.25 mm diameter) mounted on a slow-speed dental implant drill. Another bone drill (diameter 3 mm) secured to the central guide hole was used to remove the central bone and expose the bone marrow. The scaffolds (as described in Examples 8 and 9) were then placed in the marrow as follows: one leg having one control standard TiO2 (T) and one empty defect (SHAM), the other leg Have two fluoride-doped scaffolds (TOS or THF) to have internal control standards in the same animal. The TOS and THF samples were not placed together. All grinding of the bones was done with a sufficient saline solution wash. Coin-shaped titanium implants (6.25 mm diameter and 1.95 mm diameter) coated with a Teflon cap held in two titanium screws (Medicon® CMS, Germany) and a preformed titanium maxillofacial bone plate (Medicon® CMS, Germany) ) Was placed in the cortical bone as a stabilizing device for the scaffold. Flexible tissue was repositioned and closed with Dexon®II, (Sherwood-Davis & Geek, UK). After surgery, each animal was subcutaneously injected with warmed 20 ml NaCl and subcutaneously injected with 0.02 - 0.05 mg / kg buprenorphine (Temgesic ®, Reckitt & Colman, England). A second subcutaneous injection of Temgesic 0.05 mg / kg subcutaneous injection was also given at least 3 hours after the first / previous Temgesic injection. Health status was monitored during the study and the surgical site was carefully examined daily until wound healing was complete. After a healing period of 8 weeks, rabbits were intravenously injected with 1.0 ml of fluianzol / fentanyl (Hypnorm®, Janssen, Belgium) followed by 1 ml / kg intravenous injection of pentobarbital (Mebumal®, Rikshospitalets Apotek, Norway) Euthanasia. Immediately after euthanasia, incisions were made through soft tissue in the tibia. The titanium plate covered with the implants was exposed and allowed to stand. A hole was made in the center of the PTFE cap with a hollow needle and pressurized air was applied to remove the cap and expose the back portion of the titanium implant.

All bones were placed in a Skyscan 1072 (Skyscan, Aartselaar, Belgium) desktop x-ray CT (Sigma) at 7 탆 voxel resolution (50 x magnification), X-ray tube current 173 ㎂ and a voltage of 60 kV with a 0.5 mm aluminum / Scanners were used for analysis. The specimen was mounted vertically on a plastic support and rotated 180 DEG about the long axis (z-axis) of the sample. The four absorbed images were recorded each time they were rotated by 0.400 degrees. These projection random images were used for standard cone-beam reconstruction to generate a series of 1024 8-bit axial slices, each with 1024 x 1024 pixels, with Z-dimensional spacing such as the inter-slice pixel spacing. The resulting 3-D data set was therefore isotropic with a voxel spacing of 7 μm across the entire 1024 ³ spatial range. All scaffolds were aligned on both horizontal and vertical planes. 3-D reconstructions of internal pore shapes were performed using these axial bitmap images and analyzed by CTan (Skyscan, Aartselaar, Belgium). At this time, the gray scale threshold value was set to 21 to 90. This threshold filtered the titanium oxide scaffold. Therefore, analysis of the internal pore shape of the strut bone was performed together with the scaffold structure as an open space. The area of interest was chosen as the cylinder (5 mm diameter and 10 mm length) inside the trough. The shape of the cylinder was constant for the analysis of all samples. Data were later processed with statistical software (SPSS v15 for Windows, US).

result

Titanium oxide scaffolds, titanium oxide scaffolds with low fluoride and titanium oxide scaffolds with intermediate fluoride contents all increased significantly (p < 0.001) within the scaffold compared to the control standard (sham) (Fig. 4).

There was a significant difference in bone quality (p <0.001) within the scaffold compared to the control standard, but also a significant difference from the scaffold with an intermediate content of fluoride compared to the pure titanium oxide scaffold (P < 0.05) (Fig. 5), indicating fluoride stimulated bone maturation and growth.

Example 11: Preparation of slurry Titanium oxide and aluminum oxide complex

Materials and Methods:

Titanium oxide and aluminum oxide powders were dried at a ratio of 80 wt.% TiO ₂ (Pharma, Sachtleben, Germany) and 20 wt.% Al ₂ O ₃ (No. 713-40 II / 973, Nabaltec, Germany) Lt; / RTI &gt; The mixture was homogenized thoroughly in the mortar and mixed with 24 ml deionized water, 2.0 g 5% methylcellulose solution (Tylopur C 30, Hoechst, Germany) as binder and 0.2 g polycarboxylic acid (Dolapix CE 64, Zschimmer & Schwarz, Germany) And dispersed in a liquid. Next, the slurry was homogenized with an electric stirrer for 5 minutes. To achieve the optimum particle size, the ceramic slurry was dispersed in a high speed dissolution apparatus (Dispermat Ca-40, VMA-Getzmann GmbH, Reichshof, Germany) with a custom titanium rotary blade. The porous structure and sintering were carried out as described in Examples 8 and 9. In addition to the weight percentages described, 30 wt.%, 40 wt.% And 50 wt.% Of Al ₂ O ₃ were also tried.

result:

The mixture ratio of 80 wt.% TiO ₂ and 20 wt.% Al ₂ O ₃ provided the most stable solid body. Other materials are well dispersed, which can be viewed as an EDX image (Figure 6). The hardness of the samples (according to the Vickers test) is shown in FIG.

Example 12: Slurry preparation Titanium oxide and aluminum oxide complex

Materials and Methods:

Titanium oxide and aluminum oxide powder were mixed with 80 wt.% TiO ₂ (Pharma, Sachtleben, Germany), and a slurry mixture was prepared using 20 wt.% Commercial ZrO ₂ powder (3 mol% Y ₂ O ₃ , Cerac Inc., Wl, USA). 100 g of powder was vigorously stirred in 150 ml of distilled water dispersed with 6 g of triethyl phosphate (TEP; (C ₂ H ₅ ) ₃ PO ₄ , Aldrich, USA) for 24 hours. As a binder, 6 g of polyvinylbutyl (PVB, Aldrich, USA) was dissolved in another beaker, followed by addition to the slurry and stirring for an additional 24 hours. The porous structure and sintering were carried out as described in Examples 8 and 9. In addition to the weight percentages described, ZrO2 powders of 40 wt.%, 60 wt.% And 80 wt.% Were also tried.

result:

The hardness of the structure increased with increasing amount of ZrO ₂ . The highest hardness was found in the sample with 80 wt% ZrO ₂ . Figure 8 shows the Vickers hardness of a mixture of ZrO ₂ and TiO ₂ composites. Microscopic images were also shown (Figure 9).

Example 13

The composition of the mixture was as described in Examples 11 and 12, but mixing was carried out in a ball mill under wet conditions.

Claims (40)

1. A metal oxide scaffold comprising titanium oxide, wherein the scaffold has a compressive strength of 0.1 to 150 MPa, the titanium oxide is contained in an amount of 40 to 100 wt% with respect to the metal oxide scaffold, Less than 10 ppm secondary; Tertiary; Or secondary and tertiary phosphates. &Lt; RTI ID = 0.0 &gt; 11. &lt; / RTI &gt; The metal oxide scaffold of claim 1, wherein the compressive strength is 5-15 MPa. The metal oxide scaffold of claim 1, wherein the metal oxide scaffold has a porosity of 40-99%. The metal oxide scaffold of claim 1, having a pore size of 10-3000 m. The metal oxide scaffold of claim 1, having a fractal dimension strand of 2.0-3.0. 2. The metal oxide scaffold of claim 1, wherein the metal oxide scaffold has an internal pore volume of 0.001-3.0 mu m &lt; ^{3 &} gt ;. The metal oxide scaffold of claim 1, wherein the pores are interconnected or partially interconnected. The metal oxide scaffold according to claim 1, further comprising at least one oxide selected from the group consisting of Zr, Hf, V, Nb, Ta and Al. The metal oxide scaffold of claim 1, wherein the titanium oxide comprises 60-90 wt% of the metal oxide present in the scaffold. 7. The composition of claim 1, wherein the fluoride is partially or fully fluorinated; fluorine; Or at least one surface covered with fluoride and fluorine. 11. The metal oxide scaffold of claim 10, wherein the fluoride is provided in a gas phase, as a vapor, or as an aqueous solution comprising HF, NaF or CaF ₂ . The metal oxide scaffold of claim 1, wherein the metal oxide scaffold has a porosity of 70-90%. The metal oxide scaffold of claim 1, wherein the titanium oxide comprises TiO ₂ . The method according to claim 1, wherein the metal oxide comprises at least one titanium oxide selected from TiO ₂ , Ti ₃ O, Ti ₂ O, Ti ₃ O ₂ , TiO, Ti ₂ O ₃ , or Ti ₃ O ₅ Metal oxide scaffold. The metal oxide scaffold of claim 1, for regenerating, healing, replacing or repairing tissue. The metal oxide scaffold of claim 1, for regenerating, healing, substituting or repairing bones. The metal oxide scaffold of claim 1, having a pore size of 20-2000 μm. The metal oxide scaffold of claim 1, wherein the metal oxide scaffold has a pore size of 30-1500 m. The metal oxide scaffold of claim 1, having a pore size of 30-700 μm. The metal oxide scaffold of claim 1, having a fractal dimension strand of 2.2-2.3. 2. The metal oxide scaffold of claim 1, having an internal pore volume of 0.8-1.2 [mu] m &lt; ^{3 &} gt ;.  22. A medical implant comprising a metal oxide scaffold according to any one of claims 1 to 21. a) preparing a slurry of a metal oxide comprising titanium oxide, b) providing a slurry of step a) to the porous polymer structure, c) allowing the slurry of step b) to solidify, d) removing the porous polymeric structure from the solidified metal oxide slurry. &lt; RTI ID = 0.0 &gt; 21. &lt; / RTI &gt; 24. The method of claim 23, wherein step d) i) sintering the porous polymeric structure together with the solidified metal oxide slurry at 500 DEG C, holding the temperature for at least 30 minutes, ii) rapidly sintering at 1500C to 1750C at 3 K / min, maintaining this temperature for at least 10 minutes, iii) rapidly cooling to room temperature at least 3 K / min. 24. The method of claim 23, wherein the metal oxide scaffold is selected from the group consisting of fluoride; fluorine; Or fluoride and fluorine. &Lt; Desc / Clms Page number 20 &gt; 26. The method of claim 25, wherein the fluoride; fluorine; Or fluoride and fluorine are provided in a gaseous phase, as a vapor, or as an aqueous solution comprising HF, NaF or CaF ₂ . 27. The method of claim 26, wherein the fluoride in the aqueous solution; fluorine; Or a fluoride and fluorine concentration of from 0.001 to 2.0 wt%. 27. The method of claim 26, wherein the fluoride in the aqueous solution; fluorine; Or a fluoride and fluorine concentration of from 0.05 to 1.0 wt%. 22. The metal oxide scaffold of any one of claims 1 to 21, for use as a medical implant. 23. A medical implant according to claim 22, for use for tissue regeneration. 23. A medical implant according to claim 22, for use for bone regeneration. 22. A metal oxide scaffold according to any one of claims 1 to 21, characterized in that the metal oxide scaffold is granulated as a bone filling material. 24. The method of claim 23, wherein the slurry in step a) comprises a fluoride ion; fluorine; Or fluoride ions and fluorine. &Lt; RTI ID = 0.0 &gt; 11. &lt; / RTI &gt; delete delete delete delete delete delete delete

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